Liver Function Tests

Moderator – Dr. Saroj JaswalSubmitted by 51-60

GROSS ANATOMY •Two lobes (falciform ligament) • Blood supply : Hepatic artery(oxygen)& Portal vein(nutrients)• veinous supply : hepatic veins , inferior vena ceva •microscopic units lobules functional units of liver •Each lobule is a hexagonal centrally located vein with portal triads

BIOCHEMICAL FUNCTIONS The liver performs four major functions : Excretion / secretion Metabolism Detoxification Storage

1. Synthetic functions : Plasma proteins – albumin(osmotic pressure) , carrier

proteins ( transport of elements Fe Cu) , globulins , coagulation factors.

Cholestrol – precursor of steroid hormones , bile acids and 7-dehydrocholestrol.

Bile acids- to absorb lipid nutrients from the gut , includes fatty acids and fat soluble vitamins .

METABOLISM Carbohydrate metabolism :Liver is an important homeostatic regulator

of blood glucose . It can either produce glucose or store glucose

. By the following processes :1. Glycogenesis – by converting glucose into

glycogen for storage .2. Glycogenolysis – ATP is required , glycogen

is again converted back into glucose .3. Gluconeogenesis – formation of glucose

from non precursor carbohydrates such as lactate , glucogenic amino acid and propinyl CoA .

AMINO-ACID METABOLISM PROTEINS get breakdown in the intestine into

amino acids and reach liver by portal vein ; utilized to form different proteins like globulin , albumin , coagulation factors …………………..

LIPID METABOLISM : Fatty acids catabolised to release CoA – used for TCA

cycle and ETC to release energy . Acetyl CoA converted to ketone bodies – acetone ,

acetoacetate and hydroxybutyric acid . Liver synthesis apoprotein part of lipoproteins –

transportation of lipids into circulation .

PROTECTIVE FUNCTION & DETOXIFICATION

Exogenous substances : It detoxify toxic substances – hydrolysis , hydroxylation ,oxidation , reduction and demethylation .

More soluble in water and easily excreted through urine .

Drugs – metabolised by P450 enzyme system of liver , convert drugs into more soluble form which in due course conjugate with sub. Like glycine , glucouronic acid to get excreted in urine .

Endogenous substances – Kupffer cells perform phagocytosis to eliminate foreign cmpds . For eg. NH3 is detoxified urea and metabolism of xenobiotics ( detoxification )

WHAT IS PURPOSE OF LFTS?

LFTs alone do not give the physician full information, but used in combination with a careful history, physical examination (particularly ultrasound and CT Scanning), can contribute to making an accurate diagnosis of the specific liver disorder.

Different tests will show abnormalities in response to liver inflammation liver injury due to drugs, alcohol, toxins, viruses Liver malfunction due to blockage of the flow of bile Liver cancers

CLASSIFICATION OF LFT’SSynthetic function of liver-a. Serum proteins – albumin,globulin,ceruloplasmin haptoglobin , alpha

fetoprotein , alpha 1 antitrypsin levelsb. Prothrombin time

Serum Enzymes(liver enzyme panel)c. Indicating hepatocellular damage- ALT , ASTd. Indicating obstruction- ALP, GGT

Based on detoxification functione. Blood ammonia f. Hippuric acid test

Hepatic excretory function-g. serum- bilirubin levels, conjugated and unconjugated h. Urine – bile pigments , bile salts and urobilinogen

TEST BASED ON SYNTHETIC

FUNCTION OF LIVER

1)SERUM ALBUMIN LEVEL Normal Value- 3.5 to 5 g/dl Albumin is major protein- responsible for OSMOTIC

pressure in vascular system Half life: 20 days, low level in all chronic diseases of

liver Normal A/G ratio is 1.2 to 1.5 :1 Reverses in case of cirrhosis due to

hypoalbuminemia and hypergammaglobulinemia.

2) SERUM GLOBULIN

• Alpha and beta globulins synthesized by liver and immunoglobulin by lymphocytes

• Normal value - 2.5 to 3.5 g/dl • Cirrhotic liver cannot clear bacteria, antibodies against intestinal

bacteria seen.• Ig A –Inc. in alcoholic liver disease

Ig M - ↑↑ Primary biliary cirrhosis..

Ig G - ↑↑ Auto immune hepatitis.

3) PROTHROMBIN TIME

Prothrombin is synthesized by liver Half life – 6 hrs so it indicate – present function of liver PT :blood test, time taken by blood to clot Normal value : 10 to 15 sec. PT is Prolonged - liver losses 80% of reserve capacity Commonly PT is used for detecting liver coagulopathies Note : Vit K deficiency Prolongs PT time But In case of LIVER diseases : PT remains prolonged

even after parenteral administration of Vit. K

4) 1-ANTITRYPSIN (AAT)

• Most abundant 1-globulin and acute phase reactant• Inactivates : serum proteases • Normal values : 90 to 200 mg/dl• It has multiple alleles , individuals possessing PiZZ allele :

deficient activity of this enzyme : Liver cirrhosis

Low levels Neonatal cholestasis, emphysema

High levels acute trauma, infection

5) HAPTOGLOBIN• Another major 2 protein synthesised in liver.• Normal values: 30- 200 mg/dl• Function – transports free Hb in the plasma to reticuloendothelial

system (RES)• Low levels – lead to ppt of free Hb in kidneys and cause damage

• Turnover rates are less than albumin – used to Identify recent changes in the liver.

Low levels Severe hepatocellular diseases (deficient synthesis), hemolytic disease(rapid degradation)

High levels Inflammatory processes , myocardial infarction

6) -FETOPROTEIN

Normal component of fetal blood., disappears after few weeks of birth.

Normal range-up to 1 year of age < 30ng/ml adults(M and non pregnant F) < 15ng/ml Tumor marker

Maternal serum AFP level Inc. in fetal open tube neural defect and dec. in foetal down syndrome

Mild Inc. Chronic hepatitis or cirrhosis

Drastic Inc. hepatocellular carcinoma, germ cell tumour and teratoma of ovary

7) CERULOPLASMIN

Synthesized by Hepatic parenchymal cells and small part by lymphocytes

Transport : Cu Normal levels: males 22- 40mg/dl

females 25-60mg/dl

pregnancy 30-120 mg/dl

Low levels

Wilson's hepatolenticular degeneration

High levels

Acute Hepatitis, hemochromatosis, obstructive biliary disease

8)TRANSTHYREITN (PRE-ALBUMIN) Produced by liver Transport thyroxine and triidothyronine Half life 2 days , hence useful parameter to assess

hepatic function early in course of liver disorders.

TEST BASED ON SERUM ENZYMES

A. Enzymes indicating Hepatocellular Damage

Alanine amino transferase (ALT) Serum Glutamate Pyruvate transaminase (SGPT) Source - liver, cardiac muscle , skeletal muscle. Increased – when cells of the liver have been inflamed or

necrosed. Normal serum level : male- 13-35 U/L female – 10-35 U/L

SGPT PLP

Abnormal levels

Only ALT are elevated in:-1. AMI rise within 6-8 hrs and remain upto 5 days

2. Pulmonary embolism

Aspartate amino transferases (AST) Serum glutamate oxaloacetate transaminase (SGOT) Source- more liver specific

Moderate 50 - 100 Chronic liver diseases,hepatitis C, NASH(non alcoholic steatohepatitis )

Very high 300-1000

Acute hepatitis , toxic or viral in origin

ASTPLP

Normal level : 8-20 U/L

Normal AST: ALT ratio – 0.8

ratio > 2 AST is higher

Alcoholic hepatitis, hepatitis with cirrhosis , NASH , erythromycin treatment

Ratio < 0.8ALT is higher

Acute hepatocellular injury, toxic exposure ,extra hepatic obstruction

elevated liver diseases, myocardial infarct, muscle disease

B. Markers of obstructive liver disease Alkaline Phosphatase (ALP) Source: liver, bone, placenta and intestine. ALP is a hydrolase enzyme responsible for removing

phosphate groups from many types of molecules, including nucleotides and proteins.

phosphomonoester ALP alcohol + phoshate ion Normal level: 40-125 U/L Levels are significantly higher in children and pregnant

women.

Moderate increase

2-3 times Hepatic diseases includes infective hepatitis,hepatocellular carcinoma

Very high levels 10-12 times Obstructive jaundice (gall stones)

Drastically high levels

More than 12 times

Not related to liver disease but Bones disease such as rickets, osteomalacia

ISOENZYMES OF ALP Iso enzyme location Increased in

Alpha 1 ALP Epithelial cells of biliary canaliculi

Obstructive jaundice

Alpha 2 heat labile ALP

Hepatic cells

Alpha 2 heat stable ALP

Placental origin (regan iso enzyme)

Pregnancy and inhibited by phenylalanine

Pre beta ALP Bone origin Pagets disease, rickets osteomalacia

Gamma ALP Intestinal cells Ulcerative colitis5’- NucleotidaseNormal range 2-17 U/LFunction ; hydrolysis of nucleoside 5’ phosphate estersClinical significance

Sensitive and specific for hepatobiliary disorders (HBD), obstructive biliary diseases Normal pregnancy, bone growth and bone diseases do not affect 5' NTIn pts with HBD, changes in ALP are usually followed by similar changes in 5' NT

GAMMA GLUTAMYL TRANSFERASE (GGT)1. Function – Glutathione + amino acid GGT glutamyl peptide +

cysteinylglycine regulate glutathione levels

2.Source - liver, kidney, pancreas, intestinal cells absent in bone

3.Normal serum levels : 10-30 U/L4. Diagnostic signicficance hepatic microsomal enzyme GGT elevation differentiate ALP increase

levels ,seen in biliary tract diseases In alcoholic liver disease GGT levels may be parallel to

alcohol intake , even when other LFT’s are in Normal range.diseases pancreatic disease , MI ,pulmonary disease

Drugs warfarin , antidepressants

TEST BASED ON DETOXIFICATIOMBlood ammonia level Normal level: 10-50 mcg/dl Index of urea synthesis by liver, marker of hepatic

encaphlopathy Ammonia is converted to urea by liver Increased levels 1. cirrhosis

2. portocaval anastomoses

Hippuric acid test Benzoyl glycine Reaction benzoic acid+glycine = hippuric acid Ingestion of sodium benzoate thn hourly excretion hippuric

acid is constant Decreases when the liver fails to detoxify

Oral Glucose tolerance test Glucose tolerance: ability of person to metabolise

a given load of glucose Glucose load : 75g in 250-300ml of water. Urine

samples are taken

EXCRETORY FUNCTIONS OF LIVER

TESTS BASED ON EXCRETORY FUNCTION

1.Serum bilirubin2.Urine bilirubin3.Urine and faecal urobilinogen4.Urine bile salts

SERUM BILIRUBIN

What is bilirubin?

•Bilirubin is a linear tetrapyrrole structure.

•It is the end product of heme catabolism.

•It is yellowish in colour and found in bile.

•Produced by reticuloendothelial system

Since the bilirubin is a waste product, hence

it has to be excreted from the body

For this it has to be first conjugated by the

hepatocytes to make it water soluble to be

removed from the body.

CONJUGATION OF BILIRUBIN

EXCRETION OF BILIRUBIN

Water soluble bilirubin Mixed with the bile

Reaches the Intestine Deconjugated by the bacterial flora

Free Bilirubin

Reduced

Urobilinogen (UBG) (Tetrapyrrole structure)

reduction of vinyl substituent groups

Stercobilinogen (SBG)

NORMAL VALUES

Total Bilirubin 0.2 to 0.8

mg/dl

Conjugated bilirubin 0 to 0.2

mg/dl

Unconjugated Bilirubin 0.2 to

0.6mg/dl

HYPERBILIRUBINEMIAS

These can be grouped in two ways1. Conjugated or Unconjugated2. Congenital and aquired

CONGENITAL HYPERBILIRUBINEMIAS

Crigler Najjar Syndrome- Deficiency of UDP glucuronyl transferase

Gilbert Syndrome-Defective uptake of bilirubin by the liver

Dubin Johnson Syndrome-Defective excretion of conjugated bilirubin

Rotor Syndrome

AQUIRED HYPERBILIRUBINEMIAS

Physiological Jaundice in new borns Breast Milk Jaundice

URINE UROBILINOGEN

Bilirubin in Urine:Normally bilirubin is absent in urine.Conjugated bilirubin being water soluble is excreted in urine in obstructive jaundice.

This can be detected by Fouchet’s test

Urine urobilinogen - normally trace amounts is present.

In obstructive jundice no urobilinogen is present in urine.

because bilirubin cannot enter intestine.

Note: Presence of bilirubin in urine and absence of urobilinogen in urine is seen in obstructive jaundice.

In hemolytic jaundice increased production of bilirubin causes increased formation of urobilinogen which appears in urine.

Note: Increased urobilinogen in urine and absence of bilirubin in urine is seen in hemolytic jaundice.

Fecal urobilinogen - Normal about 300mg.

Increased in Hemolytic jaundice in which color of feces is dark.

In Obstructive jaundice urobilinogen is not excreted through feces and the color is the feces is pale.

BILE SALTS

SYNTHESIS OF BILE SALTS

Cholesterol hydroxylated at 3/7/12 positions

Removal of 3-carbon unit, to make it 24 C

Conjugation with glycine

Secretion into intestinal canal

In the intestine, deconjugation and removal of hydroxyl groups.

Normally bile salts (sodium salts of taurocholic acid and glycocholic acid) are present in the bile; but are not seen in urine. Bile salts in urine are detected by Hay’s test.

Positive Hay’s test indicates the obstruction in the biliary passages causing regurgitation of bile salts into the systemic circulation leading to its excretion in urine.

Jaundice

• Jaundice is yellowish discoloration of the skin, sclera and mucous membranes due to hyperbilirubinemia and deposition of bile pigments .

• Equilibrium between bilirubin production and clearance is disturbed .

• Serum bilirubin level greater than 2mg/dL

• Jaundice is NOT a disease, but rather a sign that can occur in many different diseases.

What is Jaundice?

TYPES OF JAUNDICE

PRE HEPATIC HEPATIC POST HEPATIC

Excessive amount of bilirubin is presented to the liver due to excessive hemolysis

Impaired cellular uptake, defective conjugation or abnormal secretion of bilirubin by the liver cell

Impaired excretion due to mechanical obstruction to bile flow

Elevated unconjugated bilirubin in serum

Both conjugated and unconjugated bilirubin may be elevated in serum

Elevated conjugated bilirubin in serum

TYPES OF JAUNDICE

PRE HEPATIC

HEPATIC POST HEPATIC

Hemolytic Anemia

Hepatitis, cirrhosis, Crigler-Najjar Syndrome, Dubin-Johnson Syndrome, Rotor’s Syndrome

Gallstone, malignancy, inflammation

There are other types of Jaundice : Pathologic

JaundicePathologic jaundice can occur in children and adults and is diagnosed when jaundice presents a health risk. Several forms of hepatitis, cirrhosis of the liver and other liver diseases, bile duct blockage, along with infections and medications, can also cause pathological jaundice. Gilbert Syndrome Jaundice

Gilbert's syndrome is a harmless hereditary condition that results in mild jaundice. During times of illness or stress, people with Gilbert's syndrome will experience low levels of some bilirubin-processing enzymes in their livers, according to LabTestsOnline.com. Once diagnosed, Gilbert's syndrome does not require further medical treatment.

Neonatal Jaundice

•Jaundice is clinically detectable in the newborn when the serum bilirubin levels are greater than 85 μmol/L. This occurs in approximately 60% of term infants and 80% of preterm infants.

•Neonatal jaundice first becomes visible in the face and forehead. Blanching reveals the underlying colour. Jaundice then gradually becomes visible on the trunk and extremities.

Function test Pre-hepatic Jaundice Hepatic Jaundice Post-hepatic Jaundice

Total bilirubin Normal / Increased Increased

Conjugated bilirubin Normal Increased Increased

Unconjugated bilirubin Normal / Increased Increased Normal

Urobilinogen Normal / Increased Increased Decreased / Negative

Urine Color Normal Dark (urobilinogen + conjugated bilirubin)

Dark (conjugated bilirubin)

Stool Color Normal Normal/Pale Pale

Alkaline phosphatase levels

Normal

Increased

Alanine transferase and Aspartate transferase levels

Increased

Conjugated Bilirubin in Urine Not Present Present

Splenomegaly Present Present Absent

Table of diagnostic tests

VAN DEN BERGH TEST

Van den bergh test is specific for bilirubin. Normal serum gives a positive van den bergh reaction.

Principle of the reaction:REACTION

Bilirubin + diazotized sulphanilic acid

Purple coloured azobilirubin

VAN DEN BERGH TEST

Direct Positive:

conjugated bilirubin gives a purple color immediately on addition of the reagent.

Indirect Positive:Purple color develops only when the reagent and methanol are added.

Unconjugated bilirubin gives color only when methanol is added.

Hence conjugated bilirubin is also called direct bilirubin and unconjugated called indirect bilirubin

VAN DEN BERGH TEST

BiPhasic:Purple color develops immediately on addition of reagent.Addition of methanol intensifies the color.Elevation of both unconjugated and conjugated bilirubin.

VAN DEN BERGH TEST

RESULTSIndirect Positive Hemolytic jaundiceDirect Positive Obstructive

jaundiceBiphasic Hepatic

jaundice

SCHLESINGER’S TEST

The bilinogens form complexes with zinc ions which exhibit brilliant green fluorescence.

It is negative in normal urine.

EHRLICH’S TEST

Bilinogens react with Erlich’s aldehyde reagent i.e. Para dimethyl amino benzaldehyde to form red colour.

HAYS TEST

Hay's test, also known as Hay's sulphur flower test, is a chemical test used for detecting the presence of bile salts in urine

THANK YOU